1. Field of the Invention
The present invention relates to transgenic mouse models and, more particularly, to a transgenic mouse expressing human Pyrin-domain only (PYD) protein 2 (POP2).
2. Description of the Related Art
Inflammation is critical for clearing infections and responding to injury. However, excessive or prolonged inflammation contributes to irreversible tissue damage and the dysfunction of vital organs. Pro-inflammatory chemokines and cytokines, such as TNFα, IL-6, IL-8 and IL-1β, mediate inflammation. Most of these mediators are readily secreted as active form upon signal-induced synthesis, while the release of some other cytokines is tightly controlled. For example, the production of leaderless cytokines, such as IL-1 and IL-18, is controlled by several layers of enzymatic processes; (i) IL-1 or IL-18 is synthesized as a pro-form, which is cleaved by caspase-1 (a cysteine protease), (ii) the caspase-1, itself, is also synthesized as a pro-caspase-1, which is cleaved into active caspase-1 by inflammasome, and (iii) the inflammasome is a multi-protein complex structure formed by two or more proteins scattered in the cytoplasm, wherein they form a platform for self-cleaving of pro-caspase-1 into active caspase-1. Thus, the involvement of many steps in processing these cytokines highlights the importance of possible regulatory molecules that control unfettered activation and release of these cytokines as to avoid harmful effects in mammalian host.
Although the assembly of inflammasome structure is initiated by cytosolic sensors belonging to either the nod-like receptor (NLR) or the PYHIN family. NLR family proteins (e.g. NLRP3 and NLRC4) require an evolutionarily conserved Pyrin (PYD) or caspase recruitment domain (CARD), while members of the PYHIN family (e.g. AIM2) rely solely on a PYD. A homotypic PYD-PYD interaction between the sensor and apoptotic speck-like protein containing a CARD (ASC) is followed by recruitment of pro-caspase-1 via a CARD-CARD interaction.
Recently, viral and mammalian PYRIN domain-only proteins (POPs), comprised of essentially a solitary PYD, have been identified as likely regulators of inflammatory processes by inhibiting the NF-kB p65 signaling, limiting inflammasome formation, or both. Among mammalian species, POPs are evolutionarily recent, highly conserved, and appear to be restricted to higher primates, implying a unique role for these proteins in modulating the inflammatory responses. The recent identification and characterization of POP3 and an initial description of POP4 brings the number of human POP family members to four; all of which lack homologs in mice.
POP1 is expressed in human monocytes, macrophages and granulocytes, while POP2 in human testis, lymphocytes and macrophages. Moreover, knockdown of POP2 in human cells or stable expression in mouse cells has revealed the capacity of POP2 to limit the production of both TNFα and IL-1β. A recent study reported that POP3 is expressed in human monocyte and macrophages, but not B cells and T cells. POP4 exhibits a broad constitutive expression, but is induced in human macrophages. Functionally, POP1 inhibits IKKα and β, but it does not inhibit the NLRP3 inflammasome. However, POP2 impairs both NF-κB activation and NLRP3 inflammasomes; thereby limiting the production of both TNFα and IL-1β. Inhibition of NF-κB signaling by POP2 occurs at the level of NF-κB p65, likely through altering nuclear translocation of p65 and reducing the transactivation capacity of the RelA/p65 NF-κB transactivation domain 1. POP2 also reduces formation of NLRP3 inflammasomes by disrupting PYD-PYD interaction between ASC and NLRP3. The minimum peptide and specific residues of POP2 required for restricting both NF-kB activity and the NLRP3 inflammasome have been elucidated in in vitro cultured cells. Interestingly, POP3 has been shown to specifically inhibit the AIM2 inflammasome, but not that of NLRP3, while POP4 maintains a POP2-like NF-kB inhibitory capacity, but is likely not an inflammasome inhibitor.
Since mice lack the POP2 gene (as do all non-primate species), no knockout mouse model exists to elucidate the in vivo function of POP2. Thus, the exploration of POP2 function to date has been restricted to in vitro cellular models. Moreover, the creation of a mouse model using conventional approaches that results in overexpression of the relevant gene or a tissue expression pattern that is random will not be effective for replicating the expression of the gene in humans. Consequently, there is a need in the art for a mouse model that expresses POP2 in a manner consistent with the way the protein is expressed in humans and thus can be used to understand and evaluate the physiologic role of POP2 in humans.